Neuromodulation to modulate glymphatic clearance

ABSTRACT

The present invention provides materials and methods for using electrical stimulation to treat a mammal having a proteinopathy (e.g., neurodegenerative diseases) or at risk of developing a proteinopathy are provided. For example, the present invention provides materials and methods for modulating glymphatic clearance (e.g., enhancing glymphatic clearance) of pathogenic proteins.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. provisional application No.62/379,941, filed Aug. 26, 2016, and hereby incorporated by reference.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT

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BACKGROUND OF THE INVENTION 1. Technical Field

The present invention relates to materials and methods for usingneurostimulation to treat a mammal having a proteinopathy (e.g.,neurodegenerative diseases) or at risk of developing a proteinopathy.Specifically, the present invention provides materials and methods forusing electrical stimulation for modulating glymphatic clearance (e.g.,enhancing glymphatic clearance) of pathogenic proteins in a mammalhaving a proteinopathy or at risk of developing a proteinopathy.

2. Background Information

The glymphatic system (or glymphatic clearance pathway) is a macroscopicwaste clearance system for the vertebrate central nervous system (CNS)utilizing a unique system of perivascular tunnels formed by glial cellsto promote efficient elimination of soluble and insoluble proteins andmetabolites from the central nervous system (CNS). The pathway providesa para-arterial influx route for cerebrospinal fluid (CSF) to enter thebrain parenchyma and a clearance mechanism via convective movement ofinterstitial fluid (ISF) for extracellular solutes such as misfoldedproteins and unwanted metabolites to be removed from the brain.

It has been found that the aggregation of pathogenic proteins β-amyloid,α-synuclein, and C-tau in the brain may cause the deleterious effects ofnumerous disease and disorders such as traumatic brain injury/chronictraumatic encephalothopy, epilepsy, Alzheimer's disease, and Parkinson'sdisease. Removal of these pathogenic proteins has been found to havesubstantial therapeutic benefit, for example, in treating traumaticbrain injury/chronic traumatic encephalothopy, epilepsy, Alzheimer'sdisease, and Parkinson's disease.

SUMMARY OF THE INVENTION

Glymphatic clearance of pathogenic proteins may be enhanced by 1)modulating the interstitial volume directly to increase clearance viaconvection and 2) changing the interstitial volume between cells todecrease the resistance to glymphatic flow, indirectly increasingglymphatic flow (changing the interstitial volume manipulates theeffective distance between neural cell types, reducing the resistance toglymphatic flow between cells). The present invention recognizes thatthese mechanisms may be enhanced through the neurobiological effects ofelectrical stimulation in a patient.

The present invention provides materials and methods for usingelectrical stimulation to treat a mammal having a proteinopathy or atrisk of developing a proteinopathy under conditions wherein the severityof the proteinopathy or chance of developing a proteinopathy is reduced.For example, the present invention provides materials and methods forusing electrical stimulation to modulate glymphatic clearance (e.g.,enhancing glymphatic clearance) of pathogenic proteins. Variouselectrical stimulation therapies and/or emerging electrical stimulationtechnologies may be used.

In one embodiment, the present invention features a method of modulatingglymphatic clearance in a mammal. The method includes, or consistsessentially of, administering electrical stimulation to the mammal underconditions wherein the electrical stimulation is effective to enhanceglymphatic clearance of one or more pathogenic proteins from the centralnervous system of the mammal. The electrical stimulation to the mammalmay be under conditions wherein the electrical stimulation is effectiveto increase interstitial fluid (ISF)—cerebrospinal fluid (CSF) exchangein the mammal or to draw charged proteins toward venous return.

It is thus a feature of at least one embodiment of the invention toprovide continuous electrical stimulation delivery during patientconsciousness or unconsciousness without compromising the functions ofthe sensory nervous system.

The electrical stimulation to the mammal may be under conditions whereinthe electrical stimulation is effective to increase CSF production.

It is thus a feature of at least one embodiment of the invention toincrease the driving pressure of CSF into the perivascular space (orVirchow-Robin spaces).

The electrical stimulation to the mammal may be under conditions whereinthe electrical stimulation is effective to modulate pulsatility ofpenetrating arterial vessels.

It is thus a feature of at least one embodiment of the invention toincrease the pulsation generated by smooth muscle cells along the lengthof the pial and penetrating arteries to drive paravascular CSF influx.

The electrical stimulation to the mammal may be under conditions whereinthe electrical stimulation is effective to increase aquaporin-4 (AQP4)water channel permeability.

It is thus a feature of at least one embodiment of the invention toincrease the water movement though AQP4 water channels, increasing thepenetration of CSF into ISF.

The electrical stimulation to the mammal may be under conditions whereinthe electrical stimulation is effective to decrease the resistance ofCSF penetration in the ISF space. The electrical stimulation may beunder conditions wherein the electrical stimulation is effective todecrease intracellular fluid volume and/or increase the distance betweenneuronal/non-neuronal cells.

It is thus a feature of at least one embodiment of the invention toallow for more efficient CSF flow into the Virchow-Robin spaces.

The electrical stimulation to the mammal may be under conditions whereinthe electrical stimulation is effective to induce slow wave oscillationsin specific brain areas.

It is thus a feature of at least one embodiment of the invention toinduce conditions linked to changing interstitial volume during sleep.

Electrical stimulation to the mammal may be administered during sleep.Electrical stimulation to the mammal may be under conditions whereinglymphatic clearance during sleep is enhanced by at least 10%, 15% or20%.

It is thus a feature of at least one embodiment of the invention toadminister stimulation while the patient is unconscious and primed forglymphatic clearance. It is also a feature of at least one embodiment ofthe invention to allow the patient to function with less sleep byincreasing the protein clearance and thus decreasing the negativecognitive consequences of less sleep.

Electrical stimulation to the mammal may be administered to skinsympathetic nerves in feet during sleep.

It is thus a feature of at least one embodiment of the invention toencourage blood flow to the brain and drive clearance during periods ofincreased interstitial space during sleep, as similarly found duringfoot impact during walking or running.

Several nerves may be stimulated simultaneously to modulate sympatheticand parasympathetic arms of the autonomic nervous system in tandem.

It is thus a feature of at least one embodiment of the invention todrive clearance with synchronous stimulation.

The method may include determining whether or not the severity of asymptom is reduced using at least one of the following methods:computerized topography (CT) scan, diffuse optical imaging (DOI),event-related optical signal (EROS), magnetic resonance imaging (MRI),functional magnetic resonance imaging (fMRI), magnetoencephalography(MEG), positron emission tomography (PET), single-photon emissioncomputed tomography (SPECT), cranial ultrasound, learning tests andmemory tests, motor function testing, sensory function testing, biopsy,CSF testing, blood testing and genetic testing.

It is thus a feature of at least one embodiment of the invention totrain the patient to self-administer the electrical stimulation tomaximize response. Feedback may be in real time with feedback mechanismsdelivered to the client.

The electrical stimulation to the mammal may be under conditions whereinthe electrical stimulation to the mammal includes an implantedintravenous electrode and the electrical stimulation is effective toattract proteins to the implanted electrode subjected to the electricalstimulation. The implanted electrode may be placed within the sagittalsinus vein.

It is thus a feature of at least one embodiment of the invention to drawproteins to a waste removal location.

The implanted electrode may be inserted into a vein by a stent having aninsulated interior surrounded by external electrodes.

It is thus a feature of at least one embodiment of the invention toprevent protein build up inhibiting blood flow through the vein.

Electrical stimulation may be delivered to the implanted electrode inbiphasic pulses.

It is thus a feature of at least one embodiment of the invention tosafely generate a charge bias in the electrodes attracting the chargedproteins toward the electrodes.

The electrical stimulation may be invasive or noninvasive electricalstimulation such as vagus nerve stimulation, carotid sinus nervestimulation, transcranial direct current stimulation, transcranialmagnetic stimulation, or deep brain stimulation.

It is thus a feature of at least one embodiment of the invention tostimulate multiple areas of the brain simultaneously (i.e., the entirebrain cortex including deep brain areas). It is thus a feature of atleast one embodiment of the invention to stimulate multiple areas of thebrain simultaneously using a single input pathway connected to saidareas such as the trigeminal nerve.

In one embodiment, the present invention features a method for treatinga human having a proteinopathy. The method includes, or consistsessentially of placing an electrode over a nerve of the human having theproteinopathy and administering electrical stimulation to the electrodeunder conditions wherein electrical current is delivered to the nerveand the proteinopathy is reduced.

It is thus a feature of at least one embodiment of the invention toallow the electrical generator to be wearable by the patient and used bythe patient throughout the day without direct visual medicalsupervision.

Electrical stimulation to the mammal may be administering electricalcurrent to the trigeminal nerve.

It is thus a feature of at least one embodiment of the invention tostimulate nerves non-invasively through the patient's skin.

In one embodiment, the present invention features a method for treatinga human having a proteinopathy. The method includes, or consistsessentially of implanting an electrode within a blood vessel of thehuman having the proteinopathy and administering electrical stimulationto the electrode under conditions wherein proteins are drawn to theelectrode and the proteinopathy is reduced.

It is thus a feature of at least one embodiment of the invention toutilize the polarization of the proteins to draw proteins to a desiredlocation within the body for waste removal.

These particular objects and advantages may apply to only someembodiments falling within the claims and thus do not define the scopeof the invention.

The details of one or more embodiments of the invention are set forth inthe accompanying drawings and the description below. Other features,objects, and advantages of the invention will be apparent from thedescription and drawings, and from the claims.

BRIEF DESCRIPTION OF THE FIGURES

FIGS. 1A and 1B are charts showing blood oxygen level-dependent (BOLD)effects during vagus nerve stimulation in swine;

FIG. 2 is a schematic of an electrostimulation device being applied to apatient, specifically, transcranial direct current stimulation (tDCS)and transcranial magnetic stimulation (TMS), and an enlarged inset viewof proteins passing the blood-brain barrier in the brain as a result;and

FIG. 3 is a schematic of a venous stent with electrodes placedintravenously within a patient and electrical pulses being provided tothe stent to draw proteins to the electrodes.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

The glymphatic system is the body's waste clearance pathway for thecentral nervous system (CNS). The term “glymphatic system” was coinedbased on its similar functions to the lymphatic system and its relianceon glial cells, which surround neurons in the brain and provide supportand protection for the brain's neurons. One type of glial cell areastrocytes that ensheathe the brain's blood vessel endothelial cells andare responsible for providing biochemical support for the endothelialcells that form the blood-brain barrier, provide nutrients to thenervous tissue, maintain extracellular ion balance, and repair the brainand spinal cord following traumatic injuries.

The glymphatic system is responsible for the exchange of cerebral spinalfluid (CSF) and interstitial fluid (ISF) that is driven by arterialpulsation, respiration, slow vasomotion, and CSF pressure gradients. Inparticular, aquaporin-4 (AQP4) water channels found in the astrocyticendfeet assist with the large water fluxes into and out of the brain orspinal cord as part of the CSF-ISF exchange.

At the CSF-ISF exchange, CSF is formed in the choroid plexuses of theventricles of the brain and driven into the brain parenchyma causingconvective ISF fluid fluxes that transport waste products such asmisfolded proteins and unwanted metabolites into the ISF. Interstitialproteins are cleared into the blood through the blood-brain barrier ordirectly into the CSF via ISF bulk flow that enter the perivascularspace and travel along perivascular drainage pathways to move the ISFtoward leptomeningeal arteries at the surface of the brain and,ultimately, to cervical lymph nodes which remove the potentiallyhazardous misfolded proteins and unwanted metabolites from the brain.This exchange is modulated by arterial pulsatility and is enhancedduring sleep when the interstitial volume is increased to improve foldedproteins, waste product, and excess fluid clearance.

Increasing CSF-ISF exchange enhances clearance of neurotoxic metabolicproducts, misfolded soluble and insoluble proteins, reactivemetabolites, etc., and may be facilitated by actions at several pointsin the glymphatic system. These actions may include at least one of 1)increasing CSF production to create a driving pressure to increase flowof CSF into the Virchrow-Robin spaces, 2) dilating and/or modulatingpulsatility of the penetrating arterial vessels, 3) increasing thepermeability of AQP4 water channels expressed in astrocytic endfeet thatensheath the brain vasculature, 4) decreasing intracellular fluid volumetherefore decreasing the resistance to CSF penetration in the ISF space,for example by decreasing local norepinephrine, and/or 5) inducing slowwave oscillations in specific brain areas that are the hallmark of rapideye movement (REM) sleep. 6) Intravenous electrodes may also be used todraw proteins towards venous return. These actions will be furtherdescribed in detail below.

A method for treating a mammal (e.g., a human) having a proteinopathydescribed herein may include identifying the mammal as having aproteinopathy (e.g., a neurodegenerative disease) or as being at risk ofdeveloping a proteinopathy (e.g., a neurodegenerative disease). Anyappropriate method may be used to identify a mammal having aproteinopathy or as being at risk for developing a proteinopathy. Forexample, neuroimaging techniques (e.g., computerized topography (CT)scan, diffuse optical imaging (DOI), event-related optical signal(EROS), magnetic resonance imaging (MRI), functional magnetic resonanceimaging (fMRI), magnetoencephalography (MEG), positron emissiontomography (PET), single-photon emission computed tomography (SPECT),and cranial ultrasound), cognitive function testing (e.g., learningtests and memory tests), motor function testing, sensory functiontesting, biopsy, CSF testing, blood testing and/or genetic testing maybe used to identify a human or other mammal having a proteinopathy.Mammals who are at risk of developing a proteinopathy may be identifiedby genetic screening or through concomitant risk factors (hypertension,sleep apnea, etc.).

Any type of mammal having a proteinopathy (e.g., a neurodegenerativedisease) or at risk for developing a proteinopathy (e.g., aneurodegenerative disease) may be treated as described herein. Forexample, humans and other primates such as monkeys having aggregates ofproteins may be treated with electrical stimulation. In some cases,dogs, cats, horses, cows, pigs, sheep, rabbits, mice, and rats may betreated with electrical stimulation, as described herein.

When treating a proteinopathy or risk of proteinopathy in a patient asdescribed herein, the proteinopathy may be a proteinopathy associatedwith aggregation, misfolding, and/or neurotoxic accumulation of anypathogenic protein in the CNS. Examples of proteinopathy include, forexample, traumatic brain injury (TBI), epilepsy, Alzheimer's disease(AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE),amyotrophic lateral sclerosis (ALS), Huntington's disease, Lewy BodyDisease (LBD), Batten disease, frontotemporal dementia (FTD), inclusionbody myopathy (IBM), and Paget's disease of bone (PDB). A pathogenicprotein whose aggregation and/or misfolding is associated with aproteinopathy may be a wild type protein or a mutated protein. Apathogenic protein whose aggregation, misfolding, and/or neurotoxicaccumulation is associated with a proteinopathy may be a soluble proteinor an insoluble protein. Examples of pathogenic proteins whoseaggregation, misfolding, and/or neurotoxic accumulation is associatedwith a proteinopathy include, without limitation, amyloid β (Aβ),apolipoprotein E (APOE), α-synuclein (α-syn), DJ-1, LRRK2, PINK1/PARKIN,tau, C-tau), Huntingtin protein, superoxide dismutase 1 (SOD1), TARDNA-binding protein 43 (TDP-43), FUS, progranulin, and SCN1A. In somecases, a pathogenic protein whose aggregation, misfolding, and/orneurotoxic accumulation is associated with a proteinopathy may be amutated protein (e.g., mutant Huntingtin protein (mHtt), SOD1 mutantG93A). In some embodiments, the proteinopathy treated as describedherein may be amyotrophic lateral sclerosis (ALS) associated withaggregation of wild type and/or mutant forms of SOD1, TDP-43, and/orFUS. In some embodiments, the proteinopathy treated as described hereinmay be Alzheimer's disease associated with aggregation of misfolded Aβand/or associated with aggregation of tau. In some embodiments, theproteinopathy treated as described herein may be Parkinson's diseaseassociated with aggregation of α-syn. In some embodiments, theproteinopathy treated as described herein may be Huntington's diseaseassociated with aggregation of mHtt.

The materials and methods provided herein may be used to reduce theseverity of a proteinopathy (e.g., a neurodegenerative disease). Theseverity of a neurodegenerative disease may be evaluated by examiningsymptoms of neurodegenerative diseases such as, without limitation, lossof neurons, cognitive decline, motor impairments, sensory loss, memoryloss, and/or memory dysfunction. The materials and methods providedherein may also be used to as a prophylactic treatment in individualspredisposed to a proteinopathy.

The present invention may also be used to improve the condition ofmammals with sleep deprivation by using electrical stimulation as asleep enhancement, improving the clearance of proteins by at least 10%,15% or 20% in a patient during sleep, and thus, minimizing the amount ofsleep needed by the mammal before the effects of cognitive decline areseen in the patient.

Once identified as having a proteinopathy (e.g., a neurodegenerativedisease) or as being at risk for developing a proteinopathy, the mammalmay be administered or instructed to self-administer electricalstimulation.

Methods for modulating glymphatic clearance described herein may includeany appropriate form of electrical stimulation. Examples of electricalstimulation that may be used as described herein include, withoutlimitation, peripheral nerve stimulation (e.g., vagus nerve stimulationand/or carotid sinus nerve stimulation), transcranial direct currentstimulation (tDCS), deep brain stimulation (DBS), cortical stimulation,spinal cord stimulation (SCS), transcranial magnetic stimulation (TMS),focused ultrasound, infrared stimulation, direct simulation of nervesusing a light source, genetic modification to enhance sensitivity andspecificity of the nerve to stimulation with a light source(optogenetics), and use of intravascular electrodes.

The electrical stimulation may be used to electrically stimulate anyappropriate portion of the nervous system including, without limitation,the CNS (e.g., brain, spinal cord, retina, optic nerve, olfactorynerves, olfactory epithelium, ventricles, and choroid plexus), theperipheral nervous system (PNS) (e.g., vagus nerves, carotid sinusnerves, aortic nerves, accessory nerves, and spinal nerves (e.g.,cervical, brachial, and lumbosacral)), sympathetic nervous system, andparasympathetic nervous system. The present invention also provides skinsympathetic nerve stimulation in feet during sleep to encourage bloodflow to the brain (pressure waves are sent through the arteries tomodify and increase the supply of blood to the brain) and driveglymphatic clearance during periods of increased interstitial space.

The electrical stimulation may be used to electrically stimulate one ormore nerves. In some cases, where the electrical stimulationelectrically stimulates more than one nerve, the nerves may be locatedindependently or in a ganglion. Several nerves may be stimulatedsimultaneously to modulate sympathetic and parasympathetic arms of theautonomic nervous system in tandem to drive clearance with synchronousstimulation. In some cases, the electrical stimulation may be used in aclosed-loop fashion by way of combining any form of electricalstimulation mentioned with a sensing modality that may includeelectrophysiological and/or CSF flow rate measurement. Additionally,non-invasive measurements of autonomic nerve activity may be performed,including, blood pressure, galvanic skin response, heart rate, andrespiration variability.

In one embodiment, the electrical stimulation may be used to deliver anelectrical current to target the CNS. Electrical stimulation of thesetarget areas may be facilitated by any electrical current deliverymethod, for example, tDCS of the brain, TMS of the brain, ultrasoundstimulation of the brain, DBS, epidural/subdural and subcorticalelectrode stimulation of the brain, epidural electrodes to stimulate theautonomic system at the spinal cord, and transcutaneous SCS of theautonomic nerves at the spinal cord.

In one embodiment, the electrical stimulation may be used to deliver anelectrical current to target a division of the PNS such as the autonomicnerves and may be directed specifically to, for example, the auricularvagus nerve, trigeminal nerve, facial nerve, lingual nerve, cervicalvagus nerve, carotid sinus nerve, aortic depressor nerve, baroreceptorson carotid sinus bulb, and/or superior cervical ganglia. Stimulation ofthese target nerves may be facilitated by any commercially availableelectrical current delivery device, for example, CVRx (minimallyinvasive electrode for carotid baroreceptors), Neurosigma (non-invasivestimulation of the trigeminal nerve), LivaNova (implantable bipolar cuffto stimulate cervical vagus), and Gammacore (non-invasive stimulation ofthe cervical vagus).

In one embodiment, the electrical stimulation may be used to deliver anelectrical current to electrodes implanted intravascularly in a patientby injecting the patient using a needle delivery system. Implantation ofelectrodes may be facilitated by any commercially available electrodeimplantation and monitoring/communication system, for example, StimWaveSystem (for spinal cord stimulation) and Alfred Mann BION stimulator.

Effective levels of electrical stimulation may vary depending on theseverity of the proteinopathy (e.g., a neurodegenerative disease), typeof stimulation, the age and general health condition of the subject,excipient usage, the possibility of co-usage with other therapeutictreatments such as use of other agents, and the judgment of the treatingphysician.

An effective electrical stimulation voltage may be any voltage thatreduces the severity of a proteinopathy (e.g., a neurodegenerativedisease) being treated without producing significant toxicity to themammal. In some cases, an effective PNS electrical stimulation voltage(V) may be from about 0.1 V to about 20 V (e.g., about 0.5 V to about 18V, about 1 V to about 15 V, about 2 V to about 12 V, about 3 V to about10 V, or about 4 V to about 7 V). For example, an effective electricalstimulation voltage may be about 5 V. In some cases, an effective PNSelectrical stimulation current (Amps) may be from about 0.1 mA to about206 mA (e.g., about 0.5 mA to about 150 mA, about 1 mA to about 125 mA,about 1.5 mA to about 100 mA, about 2 mA to about 75 mA, about 2.5 mA toabout 50 mA, or about 3.0 mA to about 25 mA). For example, an effectiveelectrical stimulation current may be about 3.5 mA. The effectiveelectrical stimulation voltage may remain constant or may be adjusted asa sliding scale or be variable depending on the mammal's response totreatment. Various factors may influence the actual effective electricalstimulation voltage used for a particular application. For example, thefrequency of administration, duration of treatment, use of multipletreatment agents, type of neurostimulation, and severity of theproteinopathy (e.g., a neurodegenerative disease) may require anincrease or decrease in the actual effective electrical stimulationvoltage administered.

Electrical stimulation may be administered in a continuous delivery or apulsed delivery. In some cases, a PNS pulsed electrical stimulation mayhave a pulse rate of about 1 Hz to about 200 Hz (e.g., about 2 Hz toabout 150 Hz, about 5 Hz to about 100 Hz, about 8 Hz to about 75 Hz,about 10 Hz to about 50 Hz, or about 15 Hz to about 25 Hz). For example,a pulsed electrical stimulation may have a pulse rate of about 20 Hz. Apulsed electrical stimulation may maintain a constant pulse rate or apulse rate may be adjusted as a sliding scale or variable pulse ratesdepending on the mammal's response to treatment. Various factors mayinfluence the actual pulse rate used for a particular application. Forexample, the frequency of administration, duration of treatment, use ofmultiple treatment agents, type of electrical stimulation, and severityof the proteinopathy (e.g., a neurodegenerative disease) may require anincrease or decrease in the actual electrical stimulation pulse rateadministered.

The frequency of administration may be any frequency that reduces theseverity of a proteinopathy (e.g., a neurodegenerative disease) to betreated without producing significant toxicity to the mammal. Forexample, the frequency of administration may be from about once a weekto about three times a day, from about twice a month to about six timesa day, or from about twice a week to about once a day. The frequency ofadministration may remain constant or may be variable during theduration of treatment. A course of treatment with electrical stimulationmay include rest periods. For example, electrical stimulation may beadministered daily over a two-week period followed by a two-week restperiod, and such a regimen may be repeated multiple times. As with theeffective electrical stimulation voltage and electrical stimulationpulse rate, various factors may influence the actual frequency ofadministration used for a particular application. For example, theeffective amount, duration of treatment, use of multiple treatmentagents, route of administration, and severity of the proteinopathy(e.g., a neurodegenerative disease) may require an increase or decreasein administration frequency.

An effective duration for administering electrical stimulation may beany duration that reduces the severity of a proteinopathy (e.g., aneurodegenerative disease) to be treated without producing significanttoxicity to the mammal. For example, the effective duration may varyfrom several days to several weeks, months, or years. In some cases, theeffective duration for the treatment of a proteinopathy may range induration from about one month to about 10 years. Multiple factors mayinfluence the actual effective duration used for a particular treatment.For example, an effective duration may vary with the frequency ofadministration, effective amount, use of multiple treatment agents,route of administration, and severity of the proteinopathy beingtreated.

In some cases, electrical stimulation may be administered to a mammalhaving a proteinopathy (e.g., a neurodegenerative disease) or at risk ofdeveloping a proteinopathy (e.g., a neurodegenerative disease) as acombination therapy with one or more additional agents/therapies used totreat a proteinopathy (e.g., a neurodegenerative disease) and/or one ormore additional agents/therapies used to increase ISF-CSF exchange. Forexample, a combination therapy used to treat a mammal having aproteinopathy (e.g., a neurodegenerative disease) may includeadministering to the mammal (e.g., a human) electrical stimulation andone or more treatments such as medication (e.g., latrepirdine, riluzole,donepezil, galantamine, memantine, rivastigmine, exelon, and/or L-dopa),protein therapy (e.g., protein degradation therapy), and/orimmunotherapy (e.g., active vaccination and/or passive vaccination). Inembodiments where electrical stimulation is administered in combinationwith one or more additional agents used to treat a proteinopathy (e.g.,a neurodegenerative disease), the one or more additional agents may beadministered at the same time or independently. For example, theelectrical stimulation may be administered first, and the one or moreadditional agents administered second, or vice versa. In someembodiments, focused ultrasound may be used to break-up aggregations ofproteins into smaller aggregates/individual proteins, to increasemobility in conjunction with electrical stimulation to enhanceclearance.

The materials and methods provided herein may be used for usingelectrical stimulation to treat a mammal having a proteinopathy (e.g., adisease or disorder, such as a neurodegenerative disease, associatedwith protein aggregation, protein misfolding, and/or neurotoxic proteinaccumulation in the CNS). In some cases, electrical stimulation may beused to reduce the severity of a proteinopathy. In some cases,electrical stimulation may be used to modulate glymphatic clearance(e.g., enhancing glymphatic clearance) of pathogenic proteins from theCNS. In some cases, electrical stimulation may be used to enhanceclearance of one or more pathogenic proteins (e.g., aggregatedpathogenic proteins) associated with a proteinopathy from the CNS. Insome cases, electrical stimulation may be used to increase the volume ofISF and/or increase the effective distance between neurons therebylimiting ephaptic coupling implicated in abnormal circuit behaviorswhich behaviors that are the hallmark of multiple disorders of thenervous system (for example, epilepsy and Parkinson's disease). In somecases, electrical stimulation may be used to increase mitochondrialbiogenesis in the CNS to 1) improve degradation of misfolded proteins,2) clearance of misfolded proteins from the CNS, and 3) decrease thepro-inflammatory protein expression (i.e. TNF-alpha, and, associatedwith expression of misfolded proteins.

In some cases, electrical stimulation may be used to increaseinterstitial fluid (ISF)—cerebrospinal fluid (CSF) exchange. ISF-CSFexchange may be facilitated by, for example, 1) increasing CSFproduction, 2) modulating pulsatility of the penetrating arterialvessels, 3) increasing the permeability of AQP4 water channels, 4)decreasing intracellular fluid volume, and 5) inducing slow waveoscillations in specific brain areas (e.g., areas associated with REMsleep). For example, ISF-CSF exchange may be increased using sympatheticinhibition (e.g., sympathectomy), modulating adenosine and/orepinephrine/norepinephrine release, increasing interstitial space and/orvolume, activating locus coeruleus, activating choroid plexus ependymalcells, and/or activating astrocytes and/or AQP4 channels at theneurovascular junction. Electrical stimulation may also be used to 6)directly attract proteins to an intravenous electrode via application ofa stimulus waveform. Each of the possible conditions facilitatingglymphatic clearance is described in more detail below.

1. Increasing CSF Production to Create a Driving Pressure to IncreaseFlow of CSF into the Virchow/Robin Spaces, Thereby Enhancing GlymphaticClearance

Glymphatic transport of the CSF along the periarterial spaces followedby convective flow through the brain parenchyma and exit of ISF alongthe perivenous space to the cervical lymph system is driven by multiplemechanisms. One mechanism is the constant production of CSF by thechoroid plexus providing a driving force to push fluid flow through theventricular system to the subarachnoid space.

Under steady state conditions, the choroid plexus is under considerablesympathetic inhibitory tone, which causes a reduction in the net rate ofCSF production. Electrical stimulation may be used to induce a nerveblock or sympathetic inhibition at the superior cervical ganglia causingan increase in CSF formation and thereby enhancing glymphatic clearance.Various electrical stimulation therapies may be used to causesympathetic inhibition and inhibit central sympathetic pathways toincrease CSF production, for example, autonomic nerve stimulation orfocused ultrasound stimulation of the vagus nerve, aortic depressornerve, and carotid sinus nerve may be used.

2. Dilating and/or Modulating Pulsatility of Penetrating ArterialVessels into the Brain

Glymphatic transport of the CSF is also driven by cerebral arterialpulsation, driving the CSF along the periarterial spaces.

Electrical stimulation may be used to dilate arterial vessels andincrease the pulsatility/pulsation of penetrating arterial vessels inthe brain. Various electrical stimulation therapies may be used to causedilation of arteries, for example, stimulation of the trigeminal nerve,facial nerve, vagus nerve, carotid sinus nerve, and/or aortic depressornerve may be used. Direct stimulation therapies, for example, directcortical stimulation, deep brain stimulation, transcranial directcurrent stimulation, transcranial magnetic stimulation, etc., may alsobe used to stimulate local areas of brain tissues. Moreover, electricalstimulation of these nerves or local areas in a temporal pattern, i.e.,interpulse intervals that do not vary as a function of time, mayselectively cause oscillations in pressure and dilation of arteries thatalso improves glymphatic clearance.

3. Increasing Permeability of AQP4 Water Channels Expressed inAstrocytic Endfeet that Ensheathe the Brain Vasculature

AQP4 water channels are essential for transporting water in and out ofthe brain parenchyma by enhancing transmembrane water flux in astrocytesand is a major clearance pathway of ISF solutes from the brain'sparenchyma.

Electrical stimulation may be used to increase the permeability of AQP4water channels. Various electrical stimulation therapies may be used toincrease the permeability of AQP4, for example, tDCS, corticalstimulation, focused ultrasound stimulation, and DBS used to stimulatethe cortex of the brain.

4. Decreasing Intracellular Fluid Volume/Increasing the Distance BetweenNeuronal/Non-Neuronal Cells Therefore Decreasing the Resistance of CSFPenetration the ISF Space

Increases in interstitial space volume reduce tissue resistance towardsconvective flow thus permitting CSF-ISF exchange.

Electrical stimulation may be used to decrease the resistance to CSFflow through the increase in interstitial space volume. Variouselectrical stimulation techniques may be used to decrease neuralactivity, which is shown to cause neurons to shrink and expel fluidcontents into the ISF due to the decreased metabolic demand, therebydecreasing the intracellular fluid volumes and increasing the distancebetween neurons. This creates larger effective distances between cellsallowing ISF to flow more freely between cells and decreasing theresistance to CSF flow.

For example, vagal nerve stimulation and transcranial magneticstimulation may be used to suppress neural activity in areas of thebrain. Moreover, modulating the temporal pattern, i.e., interpulseintervals that do not vary as a function of time, of the electricalstimulation may optimize the interstitial space volume necessary forconvective movement and clearance of misfolded proteins.

Vagal nerve stimulation may also be used to modulate levels ofnorepinephrine in locus coeruleus (LC), the principal site fornorepinephrine release in the brain. Vagal nerve stimulation may be usedto increase norepinephrine levels and decrease neural activity, causingneurons to shrink and expel their fluid contents into the extracellularvolume and increase the interstitial volume.

5. Inducing Slow Wave Oscillations in Specific Brain Areas that are theHallmark of REM Sleep

During sleep, the extracellular space expands and contracts to improveglymphatic clearance. During slow wave oscillations, ISF volumeincreases to remove waste products in a slow-moving stream ofextracellular fluid, which have been linked to improved glymphaticclearance during sleep.

Electrical stimulation may be used to induce slow wave oscillations.Electrical stimulation may be used to activate cholinergic neurons inthe pedunculopontine tegmentum and laterodorsal tegmentum during non-REMsleep to increase the number of slow wave REM episodes as determined byEEG (theta 5-9 Hz). Similarly, local tonic activation of the thalamicreticular nucleus may be used to induce slow wave activity in aspatially restricted region of the cortex and electrical stimulation ofglobus pallidus externa may be used to induce both REM and non-REMsleep.

6. Directly Attracting Proteins to an Electrode Via Application of aStimulus Waveform

During electrical stimulation, charged proteins can accumulate on thesurfaces of electrodes of opposite charge.

A non-invasive electrode system such as transcranial direct currentstimulation, or an invasive electrode system such as an intravascularlyplaced electrode or an implanted electrode may be used to draw chargedproteins, such as amyloid beta, towards the stimulated electrode andtoward venous return, thus enhancing the clearance of misfoldedproteins. For example, an electrode system may be implanted in a veinwhile biphasic pulses may be used to draw the charged proteins to theelectrode system.

While normally the accumulation of proteins can impede the flow offluids at those locations, circumferential electrode placement mayprevent such blockages.

The clearance of pathogenic proteins from the CNS may be evaluated byany appropriate method. For example, the clearance of pathogenicproteins from the CNS may be evaluated by measuring neural activity inthe CNS and/or the PNS. In certain instances, a course of treatment andthe severity of one or more symptoms related to the proteinopathy (e.g.,a neurodegenerative disease) being treated may be monitored. Anyappropriate method may be used to determine whether or not the severityof a symptom is reduced. For example, the severity of a proteinopathy(e.g., a neurodegenerative disease) may be assessed using neuroimagingtechniques (e.g., computerized topography (CT) scan, diffuse opticalimaging (DOI), event-related optical signal (EROS), magnetic resonanceimaging (MRI), magnetoencephalography (MEG), positron emissiontomography (PET), single-photon emission computed tomography (SPECT),and cranial ultrasound), cognitive function testing (e.g., learningtests and memory tests), motor function testing, sensory functiontesting, CSF testing, blood testing, and/or genetic testing at differenttime points. Real time monitoring of neural activity in the CNS and/orthe PNS may teach the patient to self-administer the stimulation tomaximize target engagement and/or treatment effectiveness, e.g., byadjusting stimulation voltage, pulse rate, frequency, duration. Multiplefeedback mechanisms may also be utilized to teach the patient toself-administer the electrode configuration to maximize targetengagement and/or treatment effectiveness.

The invention will be further described in the following examples, whichdo not limit the scope of the invention described in the claims.

Example 1: Functional Magnetic Resonance Imaging (fMRI) Blood OxygenLevel-Dependent (BOLD) Effects in the Swine Following Vagus NerveStimulation

The vagus nerve was stimulated in three large swine under the followingconditions: 4 V, 75 Hz, 200 μsec pulse width, 6 sec, n=3. Functionalmagnetic resonance imaging (fMRI) was used to measure changes in bloodoxygen level dependent (BOLD) effects in the brain before and afterelectrical stimulation in multiple brain structures (e.g., nucleusaccumbens, insula, prefrontal cortex, cerebellum). The BOLD response wasused as an indicator of hemodynamic response and a marker of neuralactivity in the brain.

Referring to FIGS. 1A and 1B, the BOLD response (shown on the y-axis 10)is shown over time (shown on the x-axis 12) with electrical stimulationperiods 14 of six seconds being administered with sixty second restperiods 16 in between and then repeated five times. As shown in FIG. 1A,the stimulation was found to produce a positive BOLD responsedemonstrating increased neural activity in some areas of the brain(e.g., nucleus accumbens and cerebellum), and when administered in atemporal pattern (e.g., six seconds stimulation followed by sixtyseconds rest and repeated five times) produces a positive BOLD responsefollowed by a negative BOLD response to affect arterial pulsatility. Asshown in FIG. 1B, the stimulation was found to produce negative BOLDresponse demonstrating decreased neural activity in other areas of thebrain (e.g., prefrontal cortex and insula), and when administered in atemporal pattern (e.g., six seconds stimulation followed by sixtyseconds rest repeated five times) produces a negative BOLD responsefollowed by positive BOLD response to similarly affect arterialpulsatility. The results demonstrate that electrical stimulation to thevagus nerve may be used to both modulate (increase and decrease) thehemodynamic response as well as modulate (increase and decrease) neuralactivity in multiple brain structures.

Example 2: Electrode Subjected to Electrical Stimulation In-Vitro withinthe Normal Parameters of Neuromodulation Therapy

An electrode was subjected to electrical stimulation in-vitro inphosphate buffered saline with bovine serum albumin (proteinconcentration standard) added. Electrical stimulation was performedunder the following conditions within the standard parameters ofelectrical stimulation therapies: 10 mA, 500 microsecond pulse width,100 Hz.

It was found that the electrical stimulation selectively attractedcharged proteins to the electrodes in comparison to unstimulatedcontrols. Bovine serum albumin was shown to aggregate on the stimulatedelectrode while unstimulated electrodes showed no aggregation ofmisfolded proteins.

Example 3: Electrical Stimulation of the V1 Branch of the TrigeminalNerve to Enhance Glymphatic Clearance

Referring to FIG. 2, stimulating the trigeminal nerve 18 to enhanceglymphatic clearance in human patients 20 may be implemented asdescribed below.

The present invention may be utilized to treat patients 20 sufferingfrom a number of proteinopathies such as Alzheimer's disease andParkinson's disease, and in patients 20 where trauma to the brain causesa deficit in clearance of metabolites and misfolded proteins that leadto secondary damage or impairs learning/memory (e.g., traumatic braininjury, stroke, etc.). The system may also be used to prophylacticallytreat patients 20 deemed at risk for proteinopathies due to high bloodpressure, genetic screening, etc. Finally, the system may be used totreat patients 20 with depression, anxiety and epilepsy by increasingthe influx of CSF into the ISF and thereby decreasing distance betweennerve cells and reducing ephaptic coupling.

Electrical stimulation may be applied to the V1 branch of the trigeminalnerve 18. The supraorbital branches of the trigeminal nerve 18 extendfrom the eye socket across the forehead 22 proximate the skin.Non-invasive electrodes 24 may be placed on the skin above thesupraorbital branches of the trigeminal nerve 18 and electrical currentdelivered to the electrodes 24 to activate the trigeminal nerve 18 ortrigeminal nerve fibers.

Trigeminal nerve 18 stimulation increases glymphatic clearance throughthree separate and distinct physiological mechanisms previouslydiscussed above. It is understood that different patients 20 may receivevarying amounts of benefit based on the optimization of stimulusparameters for a particular physiological mechanism.

The first physiological mechanism by which trigeminal nerve 18stimulation may enhance glymphatic clearance is by entrainment at gamma(30-90 Hz) or theta (4-12 Hz) rhythms typically generated through visualstimuli and transcranial magnetic stimulation, respectively. Unlike thepresentation of a flashing visual field or transcranial magneticstimulation which makes very loud noises during operation and requiresan expensive, large system, trigeminal nerve 18 stimulation may beadministered via a non-invasive adhesive bandage 26 with electrode 24contacts placed over the right and left supraorbital branches duringsleep.

A second physiological mechanism by which trigeminal nerve 18stimulation may enhance glymphatic clearance is by dilating thecerebral/pial arteries and other major vessels in the brain. Dilation ofthese arteries causes the vessel to displace CSF in the Virchow-Robinspace surrounding the vessel, forcing CSF into the ISF through AQP4channels, enhancing glymphatic flow and therefore clearance.Trigeminal/facial nerve stimulation dilates the cerebral arteries anddescending vessels. The autonomic pathways may be stimulatedintermittently to deliberately cause the dilation and then constrictionof these vessels to introduce pulsatility to enhance CSF penetrationinto the ISF.

The third physiological mechanism by which trigeminal nerve 19stimulation may enhance glymphatic clearance is by reducing corticalexcitability. Norepinephrine antagonists or other pharmacologicaltreatments to decrease the activity of neurons causes the neurons toshrink and expel their fluid contents into the extracellular volume.This in turn causes an increase in ISF volume, expanding the distancebetween cells that obstructing glymphatic flow, thereby decreasing theresistance to glymphatic flow. Methods that reduce neural excitabilityenhance glymphatic clearance by increasing glymphatic flow.

Referring to the inset of FIG. 2, these physiological mechanisms worktogether to increase the outward flow of misfolded proteins and otherwaste materials 28 from the brain 30 to the blood 32 through theblood-brain barrier 34 composed of astrocytic endfeet 36 that ensheathethe brain 30 vasculature and tight junctions 38 between endothelialcells 40 around the capillaries 42.

In one embodiment of the present invention, a pulse generator system 44may include electrodes 24 that are placed over the supraorbital branchof the trigeminal nerve 18 using multiple electrodes 24 placed on amildly adhesive strip 26 on the patient's forehead 22. In one embodimentone electrode 24 is placed over a single supraorbital branch on one sideof the head, with a large return electrode placed on the surface of theskin at a comfortable distance from the trigeminal (monopolar/unipolarconfiguration). In one embodiment, and as shown in FIG. 2, two or moreelectrodes 24 are placed over both the right and left supraorbitalbranch of the trigeminal nerve 18, and combinations of electrodes 24(bipolar and tripolar configurations, changes in electrode orientationto maximize activation) are used to optimize target engagement withoutinducing unwanted side effects (direct muscle activation,pain/irritation).

The electrodes 24 are connected to an external pulse generator 46 via aconducting wire 48, which provides the energy for stimulation. Theexternal pulse generator 46 may provide a lightweight wearable housing50 having a battery 52 that may be clipped onto the patient's clothingor adhered to the patient's body and worn underneath clothing. Theexternal pulse generator 46 may be worn by the patient 20 outside of thehospital environment, for example, at home or during normal everydayactivities. Therapy may be administered during sleep (for example,continuously for at least four hours, six hours, or eight hours), or maybe used throughout the day at various intervals as convenient for thepatient 20. The therapy may be used over the span of several days,weeks, or months as needed.

The external pulse generator is capable of delivering stimulating pulseswith pulse amplitudes up to 60 Volts/60 mAs, pulse widths from 15microseconds to 10000 microseconds, and pulse frequencies ranging from0.1 Hz to 50 kHz. The extended voltage range may be high enough toensure stimulation to the nerve at a desired depth from the surface ofthe skin. The pulse width range allows flexibility in determiningchronaxie to separate intended effects from unwanted side effects, aswell as including the possibility of delivering 5 kHz sinusoids forintermittent bursts of up to 100 microseconds in duration. Although itis anticipated that therapeutic frequency ranges will typically be under50 Hz, an extended range out to 10 kHz may be desirable to enablehigh-frequency block of nerves responsible for side effect. In somecases, a high frequency carrier wave (nominal 5 kHz) may be utilizedintermittently at a lower frequency (nominal 40 Hz) to help penetratethrough muscle and stimulate the trigeminal nerve 18 and/or in brainareas where corresponding evoked activity is expected.

Nominal stimulation parameters intended to entrain the trigeminal nerve18 at beta/gamma (12-100 Hz) or theta frequencies (4-8 Hz) may be 24Volts at 1000 microsecond pulse widths (physiological mechanism 1).Nominal stimulation parameters intended to dilate the cerebral/pialand/or penetrating arteries may be 280 microsecond pulse widthcathodic-leading biphasic pulses at 10 Hz and 24 Volts (physiologicalmechanism 2). Temporal patterning (turning on/off to cause oscillatingdilation/constriction of the vessel) at a nominal frequency of 0.2 Hz ofthe electrical stimulus may be introduced to induced pulsatility.Nominal stimulation parameters intended to reduce cortical activity are24 Volts, 333 Hz, at 0.5 msec pulse widths (physiological mechanism 3).

Therapeutic parameters may be optimized for each of the physiologicalmechanisms outlined above, which may provide the desired trade-offbetween therapeutic effect and therapy limiting side effects (e.g.,induced muscle twitching, pain, irritation). It is anticipated that thepatient 20 will be trained to slowly increase the current amplitude ofstimulation until just before perceptible side effects to maximizetherapeutic effects. In order to prevent irreversible electrochemicalreactions and maximize target engagement with the trigeminal nerve 18,in one embodiment, the operating mode is a Lily biphasic, cathodicleading pulse. In other embodiments, anodic leading pulses andasymmetrical pulses may also be utilized to increase tolerable dosewithout side effects. In other embodiments, a sinusoid pulse may beused. Constant current pulses may be used such that the charge densitydelivered will not change as a function of impedance changes at theelectrode interface, but constant voltage pulses may also be utilized insome embodiments to limit the complexity of the pulse generator system44.

Multiple feedback mechanisms may be utilized to teach the patient 20 toself-administer the electrode configuration to maximize targetengagement with the trigeminal nerve 18 and/or maximize glymphaticclearance. In one embodiment, the clearance of gadolinium contrast isused in conjunction with magnetic resonance imaging (MRI) as a surrogatefor the movement of misfolded proteins. In another embodiment, imagingmodalities such as MRI/SPECT/PET are used to demonstrate evokedtrigeminal nerve 18 activity is being transmitted to the brain (singlephoton emission computed tomography {SPECT}, position emissiontomography {PET}). Alternatively, recording electrodes or other sensingmodalities such as optical coherence tomography (OCT) may be used torecord and optimize evoked neural activity at the trigeminal nerve 18and/or in brain areas where corresponding evoked activity is expected.

Unlike visual, auditory, and haptic stimuli which will induce neuralactivity only in the small region of sensory cortex associated withthose sensory inputs—and therefore only enhance glymphatic clearance inthese areas—the V1 branch of the trigeminal nerve 18 projects to thenucleus tractus solitaries (NTS), the locus coeruleus, reticularformation, raphe nuclei and thalamic structures, and thereafter othersensory, limbic, and other cortical and subcortical structures affectingmore areas. Also unlike visual, auditory, or haptic stimulus, electricalstimulation of the V1 branch of the trigeminal nerve 18 does notoverride existing sensory pathways that are necessary for normalfunction (e.g., vision, hearing, touch). Rather, invasive ornon-invasive electrical stimulation of the V1 branch of the trigeminalnerve 18 may be utilized during sleep and, for example, continuously formore than 8 hours.

Example 4: Transcranial Magnetic Stimulation of the Central NervousSystem to Enhance Glymphatic Clearance

Referring again to FIG. 2, an alternative embodiment of the presentinvention is shown. In this embodiment, TMS may be used to enhanceglymphatic clearance in human patients 20. A magnetic field generator orcoil 54 is placed near the head of the patient 20. The coil 54 isconnected to a pulse generator 56 that delivers electric current to thecoil 54. The electric current delivered to the coil 54 produces anelectric current to the brain of the patient 20 below the coil 54 viaelectromagnetic induction to stimulate the desired nerve or area of thebrain, for example the CNS of the patient 20.

In a similar manner as described above in Example 3, TMS may affectvarious physiological mechanisms known to enhance glymphatic clearancein the brain.

Example 5: Electrical Stimulation from a Venous Stent Placed within theSagittal Sinus Vein

Referring to FIG. 3, drawing misfolded proteins towards intravenouselectrodes and venous return to enhance glymphatic clearance in humanpatients 20 may be implemented as described below.

A vascular stent 58 may be implanted within the lumen 60 of a patient'sblood vessel 62. The vascular stent 58 has a tubular construction whichincludes external electrodes 64 surrounding an inner layer of insulation68. The external electrodes 64 are connected to an implantable pulsegenerator (IPG) 66 providing a pulse 70 (e.g., monophasic, symmetricalbiphasic, non-symmetrical biphasic) to direct current outward toward theinner walls of the blood vessel 62. The external electrodes 64 may beplaced externally to the inner layer of insulation 68 to preventaggregation of charged proteins 28 in the blood flow path through thestent 58 and preventing blood flow through the blood vessel 62 as shownby arrow 72.

In one embodiment of the present invention, the patient 20 will beassessed by anesthesia for suitability to undergo general anesthesia.Enteric-coat acetylsalisylic acid 81 mg daily and clopidogrel 75 mgdaily will be prescribed for five days before the stent placementprocedure. Patients 20 with a history of thrombophilia will beanticoagulated before the procedure and receive single agentantiplatelet therapy. The procedure is to be performed under generalanesthesia with systemic anticoagulation using intravenous heparinadministered to maintain an activated clotting time of more than twicethe normal level.

Femoral access is to be obtained and a guide catheter positioned intothe right or left jugular bulb of the patient 20. A high-flowmicrocatheter is then navigated into the superior sagittal sinus withthe support of a microwire. Venography will be performed by selectivecontrast injections through the microcatheter to aid in visualization ofplacement of the stent 58. The IPG 66 may provide stimulus energy andmay be placed in the chest of the patient 20, with a lead wire 74connecting the stent electrodes 64 to the IPG 66 held within the vein 62traversing and exiting through the subclavian before connection to theIPG 66.

Maximizing clearance of proteins is accomplished by stimulating theelectrodes 64 using constant-current, charged-balance, cathodic-leading,biphasic pulses 70 to safely generate a small DC bias on the electrodes64. Charge density will be limited to 50 micro Coulombs/cm2 to preventunwanted electrochemical reactions. Nominal stimulation parameters are100 Hz, 500 microsecond pulse width, and 10 mA, although the currentamplitude may be adjusted for the maximum tolerable level of the patient20.

The stimulated electrodes 64 will attract charged proteins within thelocal vicinity of the electrode 64 to be cleared via para-venous efflux.Increasing the clearance within the local vicinity of the electrode 64in turn creates an osmotic gradient to enhance convective clearance ofmisfolded proteins at a distance from the electrode 64.

It is to be understood that while the disclosure has been described inconjunction with the detailed description thereof, the foregoingdescription is intended to illustrate and not limit the scope of thedisclosure, which is defined by the scope of the appended claims. Otheraspects, advantages, and modifications are within the scope of thefollowing claims.

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this disclosure belongs. Methods and materials aredescribed herein for use in the present disclosure; other, suitablemethods and materials known in the art may also be used. The materials,methods, and examples are illustrative only and not intended to belimiting. All publications, patent applications, patents, sequences,database entries, and other references mentioned herein are incorporatedby reference in their entirety. In case of conflict, the presentspecification, including definitions, will control.

Certain terminology is used herein for purposes of reference only, andthus is not intended to be limiting. For example, terms such as “upper”,“lower”, “above”, and “below” refer to directions in the drawings towhich reference is made. Terms such as “front”, “back”, “rear”, “bottom”and “side”, describe the orientation of portions of the component withina consistent but arbitrary frame of reference, which is made clear byreference to the text and the associated drawings describing thecomponent under discussion. Such terminology may include the wordsspecifically mentioned above, derivatives thereof, and words of similarimport. Similarly, the terms “first”, “second” and other such numericalterms referring to structures do not imply a sequence or order unlessclearly indicated by the context. When elements are indicated to beelectrically connected, that connection may be direct or through anintervening conductive element.

When introducing elements or features of the present disclosure and theexemplary embodiments, the articles “a”, “an”, “the” and “the” areintended to mean that there are one or more of such elements orfeatures. The terms “comprising”, “including” and “having” are intendedto be inclusive and mean that there may be additional elements orfeatures other than those specifically noted. It is further to beunderstood that the method steps, processes, and operations describedherein are not to be construed as necessarily requiring theirperformance in the particular order discussed or illustrated, unlessspecifically identified as an order of performance. It is also to beunderstood that additional or alternative steps may be employed.

It is specifically intended that the present invention not be limited tothe embodiments and illustrations contained herein and the claims shouldbe understood to include modified forms of those embodiments includingportions of the embodiments and combinations of elements of differentembodiments as come within the scope of the following claims. All of thepublications described herein, including patents and non-patentpublications, are hereby incorporated herein by reference in theirentireties.

We claim:
 1. A method for treating a human having a proteinopathy byinducing glymphatic clearance, comprising: administering an electricalstimulation to a predetermined location proximate a cranial nerve of ahead of the human having the proteinopathy and at a predeterminedfrequency and predetermined temporal pattern configured to increaseglymphatic clearance characterized by increased cerebrospinal fluid flowfrom the perivascular space into the parenchyma; wherein thepredetermined temporal pattern is a repeating pattern of alternatingstimulation intervals of delivered electrical stimulation at thepredetermined frequency separated by rest intervals of no electricalstimulation; and wherein the electrical stimulation administered to thepredetermined location is configured to induce alternating constrictionand dilation of arteries to increase a driving pressure of cerebrospinalfluid flow from the perivascular space of the brain into the parenchymaand to increase removal of pathogenic proteins from the perivascularspace using rest intervals having a longer duration than the stimulationintervals.
 2. The method of claim 1, wherein administering electricalstimulation to the predetermined location is under conditions whereinthe electrical stimulation is effective to increase cerebrospinal fluid(CSF) production.
 3. The method of claim 1, wherein administeringelectrical stimulation to the predetermined location is under conditionswherein the electrical stimulation is effective to increase aquaporin-4(AQP4) water channel permeability.
 4. The method of claim 1, whereinadministering electrical stimulation to the predetermined location isunder conditions wherein the electrical stimulation is effective todecrease resistance to cerebrospinal fluid (CSF) penetration in aninterstitial fluid (ISF) space.
 5. The method of claim 4, whereinadministering electrical stimulation to the predetermined location isunder conditions wherein the electrical stimulation is effective todecrease intracellular fluid volume.
 6. The method of claim 4, whereinadministering electrical stimulation to the predetermined location isunder conditions wherein the electrical stimulation is effective toincrease a distance between respective neuronal cells or betweenrespective non-neuronal cells.
 7. The method of claim 1, whereinadministering electrical stimulation to the predetermined location isunder conditions wherein the electrical stimulation is effective toinduce slow wave oscillations in specific brain areas including at leastone of a nucleus accumbens, insula, prefrontal cortex, cerebellum,globus pallidus externa, thalamic reticular nucleus, pedunculopontinetegmentum, laterodorsal tegmentum, and cerebral cortex.
 8. The method ofclaim 7, wherein administering the electrical stimulation to thepredetermined location is administered during the human's sleep.
 9. Themethod of claim 8, wherein administering the electrical stimulation tothe predetermined location is administered under conditions whereinglymphatic clearance during sleep is enhanced by at least 10%.
 10. Themethod of claim 1, wherein the repeating temporal pattern is repeated apredetermined number of times greater than two.
 11. The method of claim1, wherein the rest interval to stimulation interval is at most a 10:1ratio.
 12. The method of claim 1, wherein the stimulation intervals andrest intervals are configured to produce alternating negative BOLDresponse and positive BOLD response, respectively, to induce respectivealternating constriction and dilation of arteries in areas of the brain.13. The method of claim 1, wherein administering the electricalstimulation to the predetermined location activates pathways to at leastone of the following brain areas: nucleus accumbens, insula, prefrontalcortex, cerebellum, globus pallidus externa, thalamic reticular nucleus,pedunculopontine tegmentum, laterodorsal tegmentum, and cerebral cortex.14. A method for treating a human having a proteinopathy by inducingglymphatic clearance, comprising: placing a non-invasive electrode at apredetermined location over a cranial nerve within the human head of thehuman having the proteinopathy; and administering an electricalstimulation to the electrode under conditions where electrical currentis delivered to the cranial nerve at a predetermined frequency andpredetermined temporal pattern configured to increase glymphaticclearance characterized by CSF flow from the perivascular space into theparenchyma and removal of pathogenic proteins from the perivascularspace; and configuring the electrical stimulation to the electrode todeliver the electrical stimulation in a repeating temporal pattern ofalternating stimulation intervals of delivered electrical stimulation atthe predetermined frequency separated by rest intervals of no electricalstimulation to the electrode; wherein the electrical stimulation to theelectrode is operable to produce alternating positive blood oxygen leveldependent (BOLD) response and negative BOLD response, respectively, andto induce alternating constriction and dilation of arteries to increasea driving pressure of cerebrospinal fluid flow from the perivascularspace of the brain into the parenchyma and to increase removal ofpathogenic proteins from the perivascular space using rest intervalshaving a longer duration than the stimulation intervals.
 15. The methodof claim 14, wherein administering the electrical stimulation to theelectrode is repeated over a span of several days.
 16. The method ofclaim 14, wherein administering the electrical stimulation to theelectrode is delivered to a trigeminal nerve of the human.